Department of Biology, University of Pennsylvania, Philadelphia, Pennsylvania, USA.
J Bacteriol. 2013 Sep;195(17):3808-18. doi: 10.1128/JB.00572-13. Epub 2013 Jun 21.
Type IV pili play important roles in a wide array of processes, including surface adhesion and twitching motility. Although archaeal genomes encode a diverse set of type IV pilus subunits, the functions for most remain unknown. We have now characterized six Haloferax volcanii pilins, PilA[1-6], each containing an identical 30-amino-acid N-terminal hydrophobic motif that is part of a larger highly conserved domain of unknown function (Duf1628). Deletion mutants lacking up to five of the six pilin genes display no significant adhesion defects; however, H. volcanii lacking all six pilins (ΔpilA[1-6]) does not adhere to glass or plastic. Consistent with these results, the expression of any one of these pilins in trans is sufficient to produce functional pili in the ΔpilA[1-6] strain. PilA1His and PilA2His only partially rescue this phenotype, whereas ΔpilA[1-6] strains expressing PilA3His or PilA4His adhere even more strongly than the parental strain. Most surprisingly, expressing either PilA5His or PilA6His in the ΔpilA[1-6] strain results in microcolony formation. A hybrid protein in which the conserved N terminus of the mature PilA1His is replaced with the corresponding N domain of FlgA1 is processed by the prepilin peptidase, but it does not assemble functional pili, leading us to conclude that Duf1628 can be annotated as the N terminus of archaeal PilA adhesion pilins. Finally, the pilin prediction program, FlaFind, which was trained primarily on archaeal flagellin sequences, was successfully refined to more accurately predict pilins based on the in vivo verification of PilA[1-6].
IV 型菌毛在多种过程中发挥重要作用,包括表面黏附作用和扭转运动。尽管古菌基因组编码了多样化的 IV 型菌毛亚基,但大多数的功能仍然未知。我们目前已经鉴定了 6 种盐沼盐杆菌的 Pilin,分别是 PilA[1-6],每个 Pilin 都含有一个相同的 30 个氨基酸的 N 端疏水性基序,该基序是一个更大的高度保守的未知功能域(Duf1628)的一部分。缺失多达 6 个 Pilin 基因中的 5 个的缺失突变体没有明显的黏附缺陷;然而,缺乏所有 6 个 Pilin(ΔpilA[1-6])的盐沼盐杆菌不能黏附在玻璃或塑料上。与这些结果一致,这些 Pilin 中的任何一个在转座子时的表达都足以在 ΔpilA[1-6]菌株中产生功能性菌毛。PilA1His 和 PilA2His 只能部分挽救这种表型,而表达 PilA3His 或 PilA4His 的 ΔpilA[1-6] 菌株的黏附能力甚至比亲本菌株更强。最令人惊讶的是,在 ΔpilA[1-6] 菌株中表达 PilA5His 或 PilA6His 会导致微菌落的形成。一种杂合蛋白,其中成熟的 PilA1His 的保守 N 端被 FlgA1 的相应 N 结构域取代,被 prepilin 肽酶加工,但它不能组装功能性菌毛,这使我们得出结论,Duf1628 可以被注释为古菌 PilA 黏附菌毛的 N 端。最后,菌毛预测程序 FlaFind,主要是基于古菌鞭毛序列进行训练的,经过改进后,可以更准确地根据 PilA[1-6]的体内验证来预测菌毛。